Localization and interaction of the proteins constituting the GAL genetic switch in Saccharomyces cerevisiae.
In Saccharomyces cerevisiae, the GAL genes encode the enzymes required for galactose metabolism. Regulation of these genes has served as the paradigm for eukaryotic transcriptional control over the last 50 years. The switch between inert and active gene expression is dependent upon three proteins--the transcriptional activator Gal4p, the inhibitor Gal80p, and the ligand sensor Gal3p. Here, we present a detailed spatial analysis of the three GAL regulatory proteins produced from their native genomic loci. Using a novel application of photobleaching, we demonstrate, for the first time, that the Gal3p ligand sensor enters the nucleus of yeast cells in the presence of galactose. Additionally, using Förster resonance energy transfer, we show that the interaction between Gal3p and Gal80p occurs throughout the yeast cell. Taken together, these data challenge existing models for the cellular localization of the regulatory proteins during the induction of GAL gene expression by galactose and suggest a mechanism for the induction of the GAL genes in which galactose-bound Gal3p moves from the cytoplasm to the nucleus to interact with the transcriptional inhibitor Gal80p.
Wightman
Raymond
R
Faculty of Life Sciences, The University of Manchester, The Michael Smith Building, Oxford Road, Manchester M13 9PT, United Kingdom.
Bell
Rachel
R
Reece
Richard J
RJ
D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins; D005690:Galactose / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; C052130:GAL4 protein, S cerevisiae; C055002:GAL80 protein, S cerevisiae; C099289:Gal3 protein, S cerevisiae; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D005690:Galactose